DESCRIPTION This is an R21 application requesting two years of support for an investigation of the novel hypothesis that H2S is a gaseous neuromodulator. Evidence is presented in preliminary results and has been published by the applicant that H2S is produced in the brain at concentrations of 50-160 micromolar and that concentrations of H2S in this range facilitate LTP and potentiate NMDA receptor-mediated responses. An enzyme that produces H2S, cystathionine beta-synthase (CBS) is found in brain at especially high levels in the hippocampus, a region that is high in NMDA receptors and is involved in LTP. From these observations, Dr. Kimura proposes the following specific aims: (1) To examine the regulation of H2S synthesis and its release using brain homogenates and slice preparations. (2) To study with electrophysiological techniques the mechanism of activation of the NMDA receptor by H2S using the Xenopus oocyte expression system as well as a hippocampal slice preparation. (3) To examine the effects of H2S on hippocampal long-term depression (LTD) by electrophysiological methods. (4) To examine the tissue localization of the synthetic enzyme CBS by using in situ hybridization and immunohistochemistry. In addition to the observations noted above H2S also appears to have actions in the periphery. Notably, H2S can cause relaxation of several smooth muscle preparations that have been precontracted by stimulation with, for example, acetylcholine. This action of H2S is seen even in smooth muscle stripped of endothelial cells. An enzyme that can synthesize H2S, cystathionine gamma-lyase (CSE) is present in many smooth muscles while CBS is present in only a few smooth muscles. Homogenates of smooth muscles produced H2S at rates comparable to brain. The applicant has also demonstrated that H2S appears to have a synergistic effect on smooth muscle with another gaseous smooth muscle relaxant, NO. Together, these observations lead the applicant to propose the following specific aims: (1) To determine whether H2S is produced by endothelial cells or by smooth muscle cells using cell culture models. (2) To examine the mechanism of smooth muscle relaxation induced by H2S, and its synergy with NO. The proposed experiments address new and potentially important physiological functions of endogenously produced H2S, and the data generated will help in the understanding of the biological basis of the neuronal modulation and the relaxation of smooth muscle induced by this gaseous messenger.